In early experiment using anti-Mouse, my phage display worked well, but over time, when I tried to use the sample, I couldn't even get the phage precipitation and blue plaques when titered. So I did spot test on E. coli, and it shows no lysis. Does it mean my phage is no longer viable? Why so? I aliquot my phage library and never use from the same tube twice. Is there any way to preserve them or prove that it's still viable?
I'm assuming you are using M13 or other filamentous phages. These tend to be very hardy so while possible, I doubt they have all died.
Is it possible that your E. coli strain has either picked up a mutation or has lost the F' episome if you are working with filamentous phages? I would check out your host strain first.
Yes, I'm currently using the M13 phage, and the E. coli strain I used is the K12 E. coli ER2738. Thank you for the insight. I might check my host strain to see if they have undergone mutations or something similar.
I was wondering the phage might have died because in previous experiment I had no trouble with generating blue plaques, and recently I couldn't, I also always make fresh culture of the E. coli.
ER2738 should be fine, just streak it out on a tetracycline plate to make sure it retains the F'. And yes wild type phage should give blue plaques on that strain. So check the Tet-R first and then if that is ok and you still see no plaques you might try to borrow some M13 from another lab and make sure the problem is not with your phage stock.
- Badges
- Science topic
- Similar topics
- Analytical Chemistry
- Extraction